This study was conducted to develop an economical optimum medium composition for the mass production of Lactobacillus plantarum and Lactobacillus reuteri, livestock probiotics. Medium ingredient factors were selected on the basis of MRS broth composition, and the 15 ingredient variables were as follows: sucrose, glucose, molasses, yeast extract, corn steep liquor, soy peptone, dipotassium phosphate, manganese chloride, magnesium chloride, tween 80, sodium chloride, sodium acetate, ammonium citrate, sodium sulphate, and ferrous sulphate. The Plackett Burman design, consisting of 20 runs, was employed for the analysis of ingredient effects on cell growth of L. plantarum and L. reuteri.